It is known that polypeptides can be expressed in a wide variety of cellular hosts. A wide variety of structural genes have been isolated from mammals and viruses, joined to transcriptional and translational initiation and termination regulatory signals from a source other than the structural gene, and introduced into hosts into which these regulatory signals are functional.
For economic reasons it would be desirable to utilize genetically engineered unicellular microorganisms to produce a wide variety of polypeptides. However, because of the inherent differences in the nature of unicellular organisms on one hand and mammalian cells on the other, the folding and processing of polypeptides in unicellular microorganisms appears to be quite different from the folding and processing that is effected in mammalian cells. As a result, mammalian polypeptides derived from unicellular microorganisms are not always properly folded or processed to provide the desired degree of biological or physiological activity in the obtained polypeptide.
To that end attempts have been made, with varying degrees of success, to express mammalian polypeptides in plants.
The first transgenic plants expressing foreign genes were tobacco plants produced by the use of Agrobacterium tumefaciens vectors as described by Horsch et al., Science, 223: 496 (1984) and De Block et al., EMBO J., 31: 681 (1984). The transgenic plants produced in these studies were resistant to an antibiotic by virtue of the introduced transgene. These first transgenic plants were produced by introducing the foreign genes into plant protoplasts, i.e., plant cells that have had their cell wall removed either by mechanical action or enzymatic digestion. Subsequent transformation methods based on regenerable explants such as leaves, stems and roots have allowed the transformation of several dicotyledonous plant species. A variety of free DNA delivery methods, including microinjection, electroporation and particle gun technologies have allowed the transformation of monocotyledonous plants, such as corn, and rice.
Expression of a multimeric protein, i.e., a protein constituted by two different polypeptide chains in significant yields has not been achieved. The factors controlling the assembly of such multimeric proteins are not well characterized except that it is known that the individual polypeptide chains must be present in sufficient quantity and in the same subcellular compartment.
The expression of a multimeric protein in plant cells requires that the genes coding for the polypeptide chains be present in the same plant cell. The probability of actually introducing both genes into the same cell is extremely remote. Even if such a single cell cotransformant is produced, the plant must then be regenerated from this single-cell cotransformant.
Multimeric proteins containing carbohydrate residues, glycopolypeptide multimers, are found in both plants and animals. Both plant and animal glycopolypeptide multimers have a common pentasaccharide core, Man.alpha.1-3(Man.alpha.1-6)Man.beta.1-4GLcNAc.beta.1-4GLcNAc-, that is directly linked to an asparagine residue present in the polypeptide. See, Kornfeld and Kornfeld, Ann. Rev. Biochem., 54: 631 (1985).
Plant and animal glycopolypeptide multimers also have outer branches of oligosaccharides directly linked to the common pentasaccharide core. Animal and plant glycopolypeptide outer branches have N-acetylglucosamine in them, while the outer branches present in yeast only contain mannose.
Plant and animal glycopolypeptide multimers contain different terminal carbohydrates that are directly linked to the outer branches of the oligosaccharides present. Animal glycopolypeptide multimers including mammalian glycopolypeptide multimers have sialic acid present as a terminal carbohydrate residue, while plant glycopolypeptide multimers do not. See Sturm et al., J. Biol. Chem., 262: 13392 (1987).
Secretory IgA is a glycopolypeptide multimer made up of two IgA molecules joining chain (J chain) and secretory component. IgA is the major class of antibody found in secretion, such as milk, saliva, tears, respiratory secretions and intestinal secretions.
Secretory IgA is resistant to denaturation caused by harsh environments. This denaturation resistance requires that the complex secretory IgA molecule containing IgA molecules, J chain and secretory component be accurately and efficiently assembled.
It has now been found that multimeric proteins can be expressed in relatively high yields in transgenic plants generated in a particular manner. In addition, the accurate and efficient expression of glycopolypeptide multimers free from sialic acid in transgenic plants has been discovered.
It has also been discovered that passive immunity can be produced in an animal by providing sialic acid-containing soluble immunoglobulin containing a variety of different antibodies having varying antigen specificities to that animal. See, Tacket et al., New England J. Med., 318: 1240 (1988) and Eibl et al., New England J. Med., 319: 1-7 (1988). To date there is no report of producing passive immunity in an animal by administering an encapsulated glycopolypeptide multimer capable of binding a preselected pathogen.
A method of producing passive immunity by administering an encapsulated, glycopolypeptide multimer that is free from sialic acid and capable of binding a preselected pathogen has been discovered.